The top layer of human skin or the epidermis is composed of many different cell types including keratinocytes, melanocytes and langerhans cells. Keratinocytes are the major cell type of the epidermis (75-80% of the total number of cells in the human epidermis). Within the epidermis the keratinocytes reside in four distinct stages of differentiation. The basal layer rests on the basal lamina separating epidermis from the dermis. These cells are large columnar rapidly proliferating cells. These basal cells migrate upward within the epidermis, initiated by the process of differentiation. The layer above the basal cells is the spinous layer. The cells in the spinous layer initiate the production of proteins characteristic of the differentiated epidermis. The granular layer, lying above the spinous layer, is characterized by electron-dense granules. This layer is responsible for the synthesis of lipid molecules required for the formation of the water impermeable barrier of the skin. The topmost layer of the skin, the stratum corneum, is formed from the granular layer by the destruction of cellular organelles. The cells in the stratum corneum, corneocytes, contain extensively cross-linked proteins, surrounded by a highly resistant cell envelope. The corneocytes are embedded in a bed of specific lipid structures (analogous to bricks on a bed of mortar) and this structure provides the protective barrier for the skin. The outermost layer of corneocytes is peeled off from the skin during the normal process of desquamation. Differentiation of the epidermal keratinocytes is the driving force for the normal desquamation process to occur. Epidermal differentiation is important for providing the essential function of the skin, namely to provide a protective barrier against the outside environment and to prevent loss of water from the body. The basal cells which have the highest rate of growth, are the least differentiated. The most differentiated cells of the stratum corneum do not have the ability to grow.
Initiation of differentiation of keratinocytes is accompanied by inhibition of their growth. The rate of synthesis of DNA determined by the incorporation of radiolabeled substrate [.sup.3 H] thymidine, is an indicator of the rate of growth of the cells. A decrease in DNA synthesis therefore indicates decrease in growth and increase in differentiation of keratinocytes.
The present invention is based, in part, on the discovery that a combination of specific active ingredients, namely an azole and short chain lipids, results in synergistic increase in differentiation, which in turn results in increased benefits to skin, such as improved conditioning, improved youthful appearance, decrease in wrinkle appearance, moisturizing, and treatment of photodamaged skin and various skin disorders, such as acne, xerosis, ichthyosis and psoriasis.
1,25-(OH).sub.2 D.sub.3 is the major biologically active metabolite of vitamin D.sub.3. 1,25-(OH).sub.2 D.sub.3 plays a central role in regulating blood calcium levels by increasing bone resorption and calcium absorption from intestine. Recent studies indicate that exogenous or endogenous 1,25-(OH).sub.2 D.sub.3 and its precursor 25-hydroxyvitamin D (25D) inhibit DNA synthesis (i.e., inhibit growth) and induce differentiation of keratinocytes. See e.g., Pillai et al. "1,25-Dihydroxy vitamin D Production and Receptor Binding in Human Keratinocytes Varies with Differentiation" The Journal of Biological Chemistry, Vol. 263, No. 11, (Apr. 15, 1988), pp. 5390-95; and Hashimoto et al., "Growth-inhibitory effects of 1,25-Dihydroxy vitamin D.sub.3 on Normal and Psoriatic Keratinocytes" British Journal of Dermatology (1990) Vol. 123, pp. 93-98.
Unfortunately, exogenously supplied (from blood and topically applied) as well as endogenously synthesized 1,25D is rapidly inactivated in epidermal cells (keratinocytes) by further hydroxylations. Inactivation of 1,25D is achieved in cells by further hydroxylations of 1,25D, beginning at positions C24 followed by C23. The present invention describes a method to inhibit this inactivation of 1,25D and thus potentlate the action of 1,25D on keratinocyte cell maturation. Inhibition of 1,25D degradation by imidazoles and potentiation of its action has been described in other cell systems; Avery S. H. et al., "Inhibition of the Hypercalcemic Action of Vitamin D with Imidazole", Endocrinology, 1971, Vol. 89, pp. 951-957; Reinhart T. A. et al., "Ketoconazole Inhibits Self-induced Metabolism of 1,25-Dihydroxyvitamin D and Amplifies Vitamin D Receptor Upregulation in Rat Osteosarcoma Cells", Arch Biochem Biophys., 1989, Vol. 272, pp. 459-465. However, the art does not teach use of azoles in combination with ceramides to enhance the maturation benefits of 1,25D in keratinocytes or in any other cell systems.
All the vitamin D hydroxylases, including the 25 and 1 hydroxylase which activate the vitamin D to the active 1,25D and the 24 and 23 hydroxylases which inactivate the 1,25D belong to a class of monooxygenases requiring a heme containing cytochrome P450 component as part of the multienzyme complex. A class of antimicotic agents containing an imidazole molecule substituted with aromatic side chains (known as "azoles") are potent inhibitors of cytochrome P450 dependent enzymes. These azoles have a wide variety of actions including inhibition of P450 enzyme systems involved in vitamin D metabolism; Avery, S. H. et al., "Inhibition of the Hypercalcemic Action of Vitamin D With Imidazole", Endocrinology, 1971, Vol. 89, pp. 951-957. The effect of imidazoles on inhibition of vitamin D hydroxylation has been therapeutically used in the control of hypervitaminosis D; (Glass, A. R. et al., "Ketoconazole Reduces Elevated Serum Levels of 1,25-Dihydroxyvitamin D in Hypercalcemic Sarcoidosis", J. Endocrinol. Invest., Vol. 13, (1990), pp. 407-413); regulation of blood calcium; (Avery, S. H. et al., "Inhibition of the Hypercalcemic Action of Vitamin D With Imidazole", Endocrinology, 1971, Vol. 89, pp. 951-957); and in enhancing the action of 1,25D; (Reinhart, T. A. et al., "Ketoconazole Inhibits Self-induced Metabolism of 1,25-Dihydroxyvitamin D and Amplifies Vitamin D Receptor Upregulation in Rat Osteosarcoma Cells", Arch Biochem Biophys., Vol. 272, (1989), pp. 459-465). Most of these studies used ketoconazole (an azole), an established inhibitor of vitamin D metabolism.
Application of azoles in skin related area is restricted to its action as an antimicrobial agent. See, Gupta, A. K. et al. "Antifungal Agents: An Overview Part I and II", J. Am. Acad. Derm., Vol. 30, (1994) pp. 677-698 and 911-933). Ketoconazole is also used commercially in shampoos for dandruff control. Recent studies also suggest the therapeutic value of azoles in inflammatory skin diseases, such as seborrheic dermatitis; Taieb, A. et al., "Topical Ketoconazole for Infantile Seborrhoeic Dermatitis", Dermatologica, Vol. 181, (1990), pp. 26-32; Faergemann, J., "Treatment of Seborrhoeic Dermatitis of the Scalp With Ketoconazole Shampoo", Acta Derm. Venerol. (Stockh) Vol. 70, (1990), pp. 171-172. Bifonazole has been shown to exert direct anti-inflammatory activity in histamine induced erythema; Petri, H. et al., "Investigations Into The Anti-inflammatory Effect of Bifonazole", In: Hay R. J. (ed). Advances in Topical antifungal therapy., Springer verlag, Berlin, (1986), pp. 120-124.
The art described above does not, however, teach the use of imidazole derivatives in regulating vitamin D metabolism or growth and differentiation in the skin or skin cell cultures. Furthermore, the art does not describe cosmetic composition containing an azole in combination with a short-chain lipid. Combinations of short chain lipids with azoles do not appear to have been described to potentlate the benefits of vitamin D in any system.
Accordingly, it is an object of the present invention to provide compositions for treatment of skin, while avoiding the disadvantages of prior art.
It is another object of the present invention to provide a skin treatment composition containing an azole in combination with a short-chain lipid.
It is another object of the invention to provide a method for treating or preventing the appearance of wrinkled, flaky, aged, photodamaged skin or skin disorders, such as acne, xerosis, ichthyosis and psoriasis.
These and other objects of the invention will become more apparent from the detailed description and examples which follow.